Synthesis, characterization, and evaluation of antimicrobial double-layer mat incorporating nisin and thyme essential oil to enhance food safety

Antimicrobial active packaging is a promising strategy to control foodborne contamination, yet the instability and rapid inactivation of natural preservatives in complex food matrices limit their practical use. Here, we fabricate and evaluate an electrospun double-layer nanobiocomposite mat engineered to protect bioactives and enable their slow release. The inner layer comprises chitosan/poly (vinyl alcohol) loaded with nisin and Shirazi thyme essential oil (CS/PVA/N/EO), while the outer layer is cellulose acetate containing thyme essential oil (CA/EO). Solutions were electrospun and characterized by SEM, FTIR, TGA, contact angle, and tensile testing; antibacterial performance was assessed by agar diffusion and a dynamic shake-flask assay. SEM revealed bead-free fibers in the inner layer with an average diameter of 144.8 ± 28.33 nm. FTIR and TGA confirmed successful incorporation of actives and thermal stability within typical processing ranges. Surface wettability was asymmetric by design: the inner layer was hydrophilic (contact angle 46.53°-72.43°) to promote release, whereas the outer CA/EO surface was hydrophobic (95.5°) to resist moisture. The double-layer exhibited adequate mechanical integrity for handling and conversion with an elongation at break of 21.66 ± 0.62%. Antimicrobial tests confirmed inhibitory activity against both Listeria monocytogenes (L. monocytogenes) and Escherichia coli (E. coli), with inhibition zones reaching 27.26 ± 1.60 mm for L. monocytogenes and 25.13 ± 1.40 mm for E. coli. In the dynamic assay, a 5 × 5 cm, 0.36 mm pad reduced L. monocytogenes by 2 logs and maintained counts below the detection limit for 48 h, whereas the control increased to 9.58 ± 0.31 log CFU/mL. These results demonstrate an effective active mat with potential application as an antimicrobial liner.